This is a second revision of a Phase I STTR proposal that received a score of 177, to develop a tissue-based genetic test for BRCA2 hereditary ovarian cancer. Six percent of unselected US ovarian cancers are BRCA2 hereditary cancers with truncating mutations in the BRCA2 gene. It is important to know which ovarian cancer patients have BRCA2 hereditary ovarian cancer because BRCA2 patients have an increased risk of breast cancer and their relatives with mutations have an increased risk of breast and ovarian cancer. These subsequent cancers would be discovered early or prevented if widespread genetic screening was available. We have developed an antibody-based tissue truncation method to identify BRCA2 hereditary cancers and have demonstrated the proof of concept for this technology. This approach visualizes protein truncation by showing with immunohistochemistry (IHC) that the N-terminus is present but the C-terminus is absent. Preliminary Data shows that 3 breast cancers with BRCA2 truncating mutations have truncated proteins by IHC while 20 sporadic breast cancers have full-length proteins detected by both N- and C-terminal antibodies. This project will fund research and development which will ultimately lead to a Phase II large clinical study of appropriate size to convince the FDA and the medical community that this test can efficiently and effectively identify patients with hereditary BRCA2 ovarian cancer. The product (test) will be an antibody-based IHC diagnostic kit for cancer tissue samples which hospital labs will use to identify hereditary cancers. The milestones for the Phase I proposal are: 1) Determine optimal tissue preparation protocols for IHC with both N- terminal and C-terminal BRCA2 antibodies. 2) Develop a quantitative scoring system for N-terminal and C- Terminal BRCA2 immunostaining on ovarian tissues with intra-assay variability of 15% and inter-assay variability of 20% or less. 3) Demonstrate that a C-terminal to N-terminal ratio for protein truncation (BRCA2 Truncation Ratio) can distinguish 20 hereditary BRCA2 ovarian cancers from 50 sporadic ovarian cases with greater than 90% sensitivity and specificity (comparing different scoring systems). 4) Develop tissue based methods for PCR-sequencing to validate results of IHC studies using the sequencing benchmark. The goal of this Phase I proposal is to develop technology for a Phase II proposal to determine if the BRCA2 truncation test can achieve 90% sensitivity and specificity in a clinical trial of 500 ovarian cancer samples (sample size to provide 30 BRCA2 hereditary cancers).Ovarian Cancer relevance: Successful completion of this Phase I research would lead to a Phase II clinical trial which could establish a new tissue-based genetic test for BRCA2 hereditary ovarian cancer based on an IHC method to visualize protein truncation. This simpler more widely applicable approach will identify many more families with hereditary breast and ovarian cancer and consequently help patients and their relatives by identifying individuals likely to develop subsequent breast or ovarian cancer which might be prevented by tamoxifen and/or screening exams and X-rays. ? ? ?
